The first protein crystal structure determined from high-resolution X-ray powder diffraction data: a variant of T3R3 human insulin-zinc complex produced by grinding

X-ray diffraction analysis of protein structure is often limited by the ava ilability of suitable crystals. However, the absence of single crystals nee d not present an insurmountable obstacle in protein crystallography any mor e than it does in materials science, where powder diffraction techniques ha ve developed to the point where complex oxide, zeolite and small organic mo lecular structures can often be solved from powder data alone. Here, that f act is demonstrated with the structure solution and refinement of a new var iant of the T3R3 Zn-human insulin complex produced by mechanical grinding o f a polycrystalline sample. High-resolution synchrotron X-ray powder diffra ction data were used to solve this crystal structure by molecular replaceme nt adapted for Rietveld refinement. A complete Rietveld refinement of the 1 630-atom protein was achieved by combining 7981 stereochemical restraints w ith a 4800-step (d(min) = 3.24 Angstrom) powder diffraction pattern and yie lded the residuals R-wp = 3.73%, R-p = 2.84%, R-F(2) = 8.25%. It was determ ined that the grinding-induced phase change is accompanied by 9.5 and 17.2 degrees rotations of the two T3R3 complexes that comprise the crystal struc ture. The material reverts over 2-3 d to recover the original T3R3 crystal structure. A Rietveld refinement of this 815-atom protein by combining 3886 stereochemical restraints with a 6000-step (d(min) = 3.06 Angstrom) powder diffraction pattern yielded the residuals R-wp = 3.46%, R-p = 2.64%, R-F(2 ) = 7.10%. The demonstrated ability to solve and refine a protein crystal s tructure from powder diffraction data suggests that this approach can be em ployed, for example, to examine structural changes in a series of protein d erivatives in which the structure of one member is known from a single-crys tal study.